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RNAi as a tool to control the sex ratio of mouse offspring by interrupting Zfx/Zfy genes in the testis

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Abstract

The objective of this study was to explore a novel method to alter the sex-ratio balance of mouse offspring by silencing the paralogous genes Zfx/Zfy (Zinc finger X/Y-chromosomal transcription factor gene) during spermatogenesis. Four recombined vectors PRZ1, PRZ2, PRZ3, and PRZ4 (RNAi-Ready-pSIREN-RetroQ-ZsGreen) were constructed for interrupting the Zfx gene. Additionally, a recombined vector Psilencer/Zfy-shRNA was constructed for interrupting the Zfy gene. Male mice were randomly divided into 8 groups, with 20 animals per group. Five groups of mice were injected with PRZ1, PRZ2, PRZ3, PRZ4, and Psilencer/Zfy-shRNA vectors, respectively. The three control groups were injected with an equal volume of physiological saline, empty RNAi-Ready-pSIREN-RetroQ-ZsGreen vector, and empty Psilencer/Zfy-shRNA vector, respectively. All groups were injected every 7 days for a total of four injections. Fourteen days after the fourth injection, 10 male mice from each group were mated individually with 10 females. Testicular tissue of 10 male mice in each group was collected, and the expression level of Zfx/Zfy mRNA was determined by qRT-PCR. Results showed that, compared with the empty RNAi-Ready-pSIREN-RetroQ-ZsGreen vector and the physiological saline group, expression of Zfx mRNA decreased significantly after injection of PRZ1 (p < 0.01), PRZ3 (p < 0.01), and PRZ4 (p < 0.01), and 78.75 ± 7.50% of the offspring were male in PRZ4 group, significantly higher than the offspring derived from the empty RNAi-Ready-pSIREN-RetroQ-ZsGreen vector and physiological saline group (p < 0.01). In the PRZ1 group, the expression of Zfx mRNA was also significantly lower (p < 0.01), but the male rate of offspring was not different (p > 0.05). Conversely, the expression of Zfy mRNA decreased significantly after injection of Psilencer/Zfy-shRNA (p < 0.01) and 31.00 ± 11.00% of the offspring were male, significantly lower than in the physiological saline group (p < 0.01). In conclusion, our findings show that RNAi-mediated disruption of Zfx/Zfy in mouse testis affected X/Y spermatogenesis. Additionally, results suggest that the paralogous genes Zfx/Zfy play an important role in the process of X and Y sperm development. The individual interference of Zfx/Zfy may predict the outcome of X and Y haploid sperms. Presented herein is an advanced method developed to control mouse X/Y spermatogenesis and sex ratio of offspring.

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Acknowledgements

We thank HaiYan Lu for assistance with data analyses, as well as HongTao Li and Hong Shen for kind assistance with electron microscopy and accurate injection respectively. We thank LetPub (http://www.letpub.com) for its linguistic assistance during the preparation of this manuscript. The funding was provided by Science and Technology supporting XinJiang from Production and Construction Corps of ShiHeZi University, China (2014AB012).

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    Authors and Affiliations

    1. College of Animal Science and Technology, Shihezi University, Xinjiang, Xinjiang Uygur Autonomous Region, China

      YongSheng Zhang, JiFeng Xi, Bin Jia, XiangZu Wang, XuHai Wang, ChaoCheng Li, YaQiang Li, XianCun Zeng, RuiWen Ying, Song Jiang & FangYuan Yuan

    2. College of Life Sciences, Shihezi University, Xinjiang, Xinjiang Uygur Autonomous Region, China

      Xin li

    3. Xinjiang Agricultural Vocational Technical College, Shihezi, China

      JiFeng Xi & XiangZu Wang

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    1. YongSheng Zhang
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    Correspondence to Bin Jia.

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    YongSheng Zhang and JiFeng Xi contributed equally to this paper and should be regarded as co-first authors.

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    Zhang, Y., Xi, J., Jia, B. et al. RNAi as a tool to control the sex ratio of mouse offspring by interrupting Zfx/Zfy genes in the testis. Mamm Genome 28, 100–105 (2017). https://doi.org/10.1007/s00335-017-9682-y

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